scholarly journals Method to obtain homozygous introgressing segments in Drosophila to identify the presence of hybrid sterility genes of the reproductive isolation

2020 ◽  
Author(s):  
Francisco García-Franco ◽  
Lilian Barandica-Cañon ◽  
Ezel Galindo-Pérez ◽  
Martha Martínez García ◽  
Blanca Chávez-Sandoval
Keyword(s):  
In Situ Hybridization ◽  
Physical Map ◽  
Hybrid Sterility ◽  
Bioinformatic Analysis ◽  
Chromosome 4 ◽  
Genomic Map ◽  
Hybrid Lines ◽  
First Time ◽  
Polythene Chromosomes

Abstract Here, we present for the first time, a method to generate homozygous segmental introgressions, by means of crosses between a pair of synmorphic species. The introgressions were monitored by the cytogenetic method of polygenic chromosome asynapses. Later the introgressions were evaluated in their capacity to produce sterility in segmental males. Also, the smallest segment with the capacity to produce sterility in segmental males was mapped by in situ hybridization of polythene chromosomes, using 8 sequences of BACs clones as probes. Finally, a bioinformatic analysis was carried out to identify the presence of particular genes. From 2 parental strains, D. buzzatii and D. koepferae, 6 simple segmental hybrid lines were generated, whose introgressing segments are distributed along chromosome 4 of these species. From the 6 simple segmental lines and by means of a new crossing strategy, the 6 respective homozygous segmental hybrid offspring were obtained, each of them carrying a specific homozygous introgression. None of the 6 heterozygous introgressions was capable of producing sterility in segmental males, while 4 of the same homozygous introgressions produced total sterility in segmental males, including in this group the two smallest introgressive segments, one of 5.03 % and the other 7.87% with respect to the total length of chromosome 4, which are located in the region F2 to F4 of the standard cytological map based on polythene chromosomes of the Drosophila Repleta group. In situ hybridization, using 8 clones from contig 1065 located along the F2 to F4 region of the physical map of D. buzzattii constructed in BACs, confirmed the precise location of the 6 clones in the chromosomal region F2 to F4 of chromosome 4 of the polygenic chromosomes of both D. buzzatii and D. mojavensis. The bioinformatic analysis of the F2 to F4 region, using the complete genetic sequence of the contig 1065 of D. buzzatti shows the presence of two predicted genes in the genomic map of D. buzzatii (g.1313.t1 and g.1314.t1), and the orthologous association of these 2 genes both with the D. moj_GI22766 gene of D. mojavensis and with the Trivet gene of D. melanogaster.

Physical mapping of barley genes using an ultrasensitive fluorescence in situ hybridization technique

Genome ◽  
2004 ◽  
Vol 47 (1) ◽  
pp. 179-189 ◽  
Author(s):  
J L Stephens ◽  
S E Brown ◽  
N L.V Lapitan ◽  
D L Knudson
Keyword(s):  
In Situ Hybridization ◽  
Hordeum Vulgare ◽  
Fluorescence In Situ Hybridization ◽  
Linkage Map ◽  
Physical Mapping ◽  
Physical Map ◽  
Primary Objective ◽  
Hybridization Technique ◽  
Hordeum Vulgare L

The primary objective of this study was to elucidate gene organization and to integrate the genetic linkage map for barley (Hordeum vulgare L.) with a physical map using ultrasensitive fluorescence in situ hybridization (FISH) techniques for detecting signals from restriction fragment length polymorphism (RFLP) clones. In the process, a single landmark plasmid, p18S5Shor, was constructed that identified and oriented all seven of the chromosome pairs. Plasmid p18S5Shor was used in all hybridizations. Fourteen cDNA probes selected from the linkage map for barley H. vulgare 'Steptoe' × H. vulgare 'Morex' (Kleinhofs et al. 1993) were mapped using an indirect tyramide signal amplification technique and assigned to a physical location on one or more chromosomes. The haploid barley genome is large and a complete physical map of the genome is not yet available; however, it was possible to integrate the linkage map and the physical locations of these cDNAs. An estimate of the ratio of base pairs to centimorgans was an average of 1.5 Mb/cM in the distal portions of the chromosome arms and 89 Mb/cM near the centromere. Furthermore, while it appears that the current linkage maps are well covered with markers along the length of each arm, the physical map showed that there are large areas of the genome that have yet to be mapped.Key words: Hordeum vulgare, barley, physical mapping, FISH, cDNA, genetics, linkage, chromosome, BACs.

Localization of the sites of synthesis and action of prolactin by immunocytochemistry and in-situ hybridization within the human utero-placental unit

1991 ◽  
Vol 7 (3) ◽  
pp. 241-247 ◽  
Author(s):  
W.-X. Wu ◽  
J. Brooks ◽  
M. R. Millar ◽  
W. L. Ledger ◽  
P. T. K. Saunders ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Culture Medium ◽  
Cdna Probe ◽  
Specific Hybridization ◽  
Decidual Cells ◽  
Its Gene ◽  
First Time ◽  
Silver Grains ◽  
Prolactin Mrna

ABSTRACT While the fetal pituitary synthesizes and releases prolactin, it is also produced within the utero-placental unit during pregnancy in women and has been localized in the amnion, chorion and decidua. However, it is not clear whether prolactin is synthesized within all these non-fetal pituitary tissues. We have investigated prolactin production and its gene expression using tissue culture, immunocytochemistry and in-situ hybridization techniques. Prolactin was immunolocalized not only in the decidua but also in amnion and trophoblast cells. In contrast, the in-situ hybridization results showed that silver grains, formed by specific hybridization of a prolactin cDNA probe to prolactin mRNA, were confined to decidual cells of early and term pregnancy. The results from tissue cultures correlated well with those of in-situ hybridization, that is that only the decidua made detectable prolactin, while it was undetectable in the culture medium from trophoblast tissue, irrespective of the stage of pregnancy. This study, for the first time, establishes that only decidualized cells are involved in biosynthesis of prolactin; other prolactin-containing cells in the amnion and trophoblast appear to sequester prolactin, possibly via receptors, suggesting that prolactin may play an important paracrine role within the amnion and syncitio- and cytotrophoblast of the utero-placental unit.

scholarly journals Fluorescence In Situ Hybridization (FISH) on Human Chromosomes Using Photoprobe Biotin-labeled Probes

2003 ◽  
Vol 51 (4) ◽  
pp. 549-551 ◽  
Author(s):  
Anja Weise ◽  
Peter Harbarth ◽  
Uwe Claussen ◽  
Thomas Liehr
Keyword(s):  
In Situ Hybridization ◽  
Fluorescence In Situ Hybridization ◽  
Yeast Artificial Chromosome ◽  
Artificial Chromosome ◽  
Dna Probes ◽  
Human Chromosomes ◽  
First Time ◽  
Established Technique ◽  
Whole Chromosome Painting

Fluorescence in situ hybridization (FISH) on human chromosomes in meta-and interphase is a well-established technique in clinical and tumor cytogenetics and for studies of evolution and interphase architecture. Many different protocols for labeling the DNA probes used for FISH have been published. Here we describe for the first time the successful use of Photoprobe biotin-labeled DNA probes in FISH experiments. Yeast artificial chromosome (YAC) and whole chromosome painting (wcp) probes were tested.

scholarly journals Catalyzed Reported Deposition-Fluorescence In Situ Hybridization Protocol To Evaluate Phagotrophy in Mixotrophic Protists

2005 ◽  
Vol 71 (11) ◽  
pp. 7321-7326 ◽  
Author(s):  
Juan M. Medina-Sánchez ◽  
Marisol Felip ◽  
Emilio O. Casamayor
Keyword(s):  
In Situ Hybridization ◽  
Fluorescence In Situ Hybridization ◽  
Signal Amplification ◽  
Cell Attachment ◽  
Cell Loss ◽  
Alexa Fluor ◽  
Hybridization Protocol ◽  
Card Fish ◽  
First Time

ABSTRACT We describe a catalyzed reported deposition-fluorescence in situ hybridization (CARD-FISH) protocol particularly suited to assess the phagotrophy of mixotrophic protists on prokaryotes, since it maintains cell and plastid integrity, avoids cell loss and egestion of prey, and allows visualization of labeled prey against plastid autofluorescence. This protocol, which includes steps such as Lugol's-formaldehyde-thiosulfate fixation, agarose cell attachment, cell wall permeabilization with lysozyme plus achromopeptidase, and signal amplification with Alexa-Fluor 488, allowed us to detect almost 100% of planktonic prokaryotes (Bacteria and Archaea) and, for the first time, to show archaeal cells ingested by mixotrophic protists.

scholarly journals EnVision+, a New Dextran Polymer-based Signal Enhancement Technique for In Situ Hybridization (ISH)

2001 ◽  
Vol 49 (9) ◽  
pp. 1067-1071 ◽  
Author(s):  
Klaus Hermann Wiedorn ◽  
Torsten Goldmann ◽  
Christof Henne ◽  
Heike Kühl ◽  
Ekkehard Vollmer
Keyword(s):  
In Situ Hybridization ◽  
Signal Amplification ◽  
Detection System ◽  
Lower Sensitivity ◽  
The Novel ◽  
Visualization System ◽  
Integrated Optical ◽  
Assay Time ◽  
First Time

Seventy paraffin-embedded cervical biopsy specimens and condylomata were tested for the presence of human papillomavirus (HPV) by conventional in situ hybridization (ISH) and ISH with subsequent signal amplification. Signal amplification was performed either by a commercial biotinyl-tyramide-based detection system [GenPoint (GP)] or by the novel two-layer dextran polymer visualization system EnVision+ (EV), in which both EV–horseradish peroxidase (EV–HRP) and EV–alkaline phosphatase (EV–AP) were applied. We could demonstrate for the first time, that EV in combination with preceding ISH results in a considerable increase in signal intensity and sensitivity without loss of specificity compared to conventional ISH. Compared to GP, EV revealed a somewhat lower sensitivity, as measured by determination of the integrated optical density (IOD) of the positively stained cells. However, EV is easier to perform, requires a shorter assay time, and does not raise the background problems that may be encountered with biotinyl–tyramide-based amplification systems. (J Histochem Cytochem 49:1067–1071, 2001)

scholarly journals Shifting into high gear: how interstitial cells of Cajal change the motility pattern of the developing intestine

2020 ◽  
Vol 319 (4) ◽  
pp. G519-G528
Author(s):  
Nicolas R. Chevalier ◽  
Yanis Ammouche ◽  
Anthony Gomis ◽  
Clémence Teyssaire ◽  
Pascal de Santa Barbara ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
In Situ Hybridization ◽  
Calcium Imaging ◽  
Interstitial Cells Of Cajal ◽  
Interstitial Cell ◽  
Sharp Transition ◽  
Anoctamin 1 ◽  
Cells Of Cajal ◽  
First Time

We reveal a sharp transition from smooth muscle to interstitial cell of Cajal (ICC)-driven motility in the chicken embryo, leading to higher-frequency, more rhythmic contractile waves. We predict the transition to happen between 12 and 14 embryonic wk in humans. We image for the first time the onset of ICC activity in an embryonic gut by calcium imaging. We show the first KIT and anoctamin-1 (ANO1) in situ hybridization micrographs in the embryonic chicken gut.

Distribution of FISH oligo-5S rDNA and oligo-(AGGGTTT)3 in Hibiscus mutabilis L.

Genome ◽  
2021 ◽  
pp. 1-10
Author(s):  
Xiaomei Luo ◽  
Zhoujian He
Keyword(s):  
Chromosome Number ◽  
Physical Map ◽  
Molecular Cytogenetics ◽  
5S Rdna ◽  
Chromosome Length ◽  
Chromosome Identification ◽  
Karyotype Asymmetry ◽  
First Time ◽  
Distribution Of Fish

Hibiscus exhibits high variation in chromosome number both within and among species. The Hibiscus mutabilis L. karyotype was analyzed in detail using fluorescence in situ hybridization (FISH) with oligonucleotide probes for (AG3T3)3 and 5S rDNA, which were tested here for the first time. In total, 90 chromosomes were counted in prometaphase and metaphase, and all exhibited similarly intense (AG3T3)3 signals at both ends. (AG3T3)3 showed little variation and thus did not allow discrimination among H. mutabilis chromosomes, but its location at both ends confirmed the integrity of each chromosome, thus contributing to accurate counting of the numerous, small chromosomes. Oligo-5S rDNA marked the proximal/distal regions of six chromosomes: weak signals on chromosomes 7 and 8, slightly stronger signals on chromosomes 15 and 16, and very strong signals on chromosomes 17 and 18. Therefore, 5S rDNA could assist in chromosome identification in H. mutabilis. Metaphase chromosome lengths ranged from 3.00 to 1.18 μm, indicating small chromosomes. The ratios of longest to shortest chromosome length in prometaphase and metaphase were 2.58 and 2.54, respectively, indicating karyotype asymmetry in H. mutabilis. These results provide an exact chromosome number and a physical map, which will be useful for genome assembly and contribute to molecular cytogenetics in the genus Hibiscus.

scholarly journals Biofilm Community Structure in Polluted Rivers: Abundance of Dominant Phylogenetic Groups over a Complete Annual Cycle

2000 ◽  
Vol 66 (7) ◽  
pp. 3078-3082 ◽  
Author(s):  
I. H. M. Br�mmer ◽  
W. Fehr ◽  
I. Wagner-D�bler
Keyword(s):  
In Situ Hybridization ◽  
Community Structure ◽  
Sampling Procedure ◽  
Single Group ◽  
Phylogenetic Groups ◽  
Polluted Rivers ◽  
River Biofilm ◽  
First Time ◽  
Biofilm Community

ABSTRACT The seasonal dynamics of river biofilm communities in two German rivers, the Elbe and one of its tributaries, the Spittelwasser, were investigated for the first time by using fluorescence in situ hybridization and a standardized biofilm sampling procedure. We show the importance of members of the β subclass of the classProteobacteria, which formed the largest single group in the massively polluted Spittelwasser at all times. Clear seasonal peaks of abundance were observed for the planctomycetes and theCytophaga-Flavobacterium cluster.

Chromatin characterization by banding techniques, in situ hybridization, and nuclear DNA content in Cicer L. (Leguminosae)

Genome ◽  
1996 ◽  
Vol 39 (2) ◽  
pp. 258-265 ◽  
Author(s):  
I. Galasso ◽  
D. Pignone ◽  
M. Frediani ◽  
M. Maggiani ◽  
R. Cremonini
Keyword(s):  
In Situ Hybridization ◽  
Dna Content ◽  
Nuclear Dna ◽  
Hybrid Sterility ◽  
Nuclear Dna Content ◽  
5S Rdna ◽  
Rrna Genes ◽  
Evolutionary Pathway ◽  
C Banding

The karyotypes of three accessions, one each from three annual species of the genus Cicer, namely Cicer arietinum, Cicer reticulation, and Cicer echinospermum, were examined and compared using C-banding, the fluorochromes chromomycin A3, DAPI, and Hoechst 33258, in situ hybridization of the 18S–5.8S–25S and 5S rDNA sequences, and silver staining. The nuclear DNA content of the three species and the amount of heterochromatin were also determined. The results suggest an evolutionary pathway in which C. reticulatum is the ancestral species from which both C. arietinum and C. echinospermum are derived with the loss of one pair of satellites; subsequently, C. echinospermum further differentiated by the accumulation of chromosomal rearrangement(s) that gave rise to a hybrid sterility barrier. Key words : Cicer, C-banding, fluorochromes, Ag staining, rRNA genes.

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